In a series of experiments, untreated and ammonium hydroxide pretreated Klenow lowland variety sw... more In a series of experiments, untreated and ammonium hydroxide pretreated Klenow lowland variety switchgrasses are converted to reducing sugars using low-frequency (20 kHz) ultrasound and commercially available cellulase enzyme. Results from experiments using untreated and pretreated switchgrasses with and without ultrasound are presented and discussed. In untreated switchgrass experiments, the combination of ultrasound and enzymes resulted in an increase of 7.5% in reducing sugars compared to experiments using just enzymes. In experiments using ammonium hydroxide pretreated switchgrass, the combination of ultrasound and enzymes resulted in an increase of 9.3% in reducing sugars compared to experiments using just enzymes. Experimental evidence indicates that there is a synergistic effect from the combination of ultrasound and enzymes which lowers the diffusion-limiting barrier to enzyme/substrate binding and results in an increase in reaction rate. Scanning electron microscopic images...
... Newman MM, Nguyen QA: Effects of temperature and moisture on dilute-acid steam explosion ... ... more ... Newman MM, Nguyen QA: Effects of temperature and moisture on dilute-acid steam explosion ... Dale BE, Ingram LO, Bothast RJ: Ethanol production from AFEX pretreated corn fiber ... Reshamwala S, Shawky BT, Dale BE: Ethanol production from enzymatic hydrolysates of AFEX ...
Use of agricultural residues for ethanol production requires pretreatment of the material to faci... more Use of agricultural residues for ethanol production requires pretreatment of the material to facilitate release of sugars. Physical–chemical pretreatment of lignocellulosic biomass can, however, give rise to side-products that may be toxic to fermenting microorganisms and hinder utilization of sugars obtained from biomass. Potentially problematic compounds include furan aldehydes formed by degradation of sugars, organic acids released from hemicellulose side-groups,
Journal of Industrial Microbiology and Biotechnology, 1999
We have developed a novel ethanologenic Escherichia coli strain FBR3 that is an efficient biocata... more We have developed a novel ethanologenic Escherichia coli strain FBR3 that is an efficient biocatalyst for converting mixed sugar streams (eg, arabinose, glucose, and xylose) into ethanol. In this report, the strain was tested for conversion of corn fiber hydrolysates into ethanol. Corn fiber hydrolysates with total sugar concentrations of 7.5% (w/v) were prepared by reacting corn fiber with dilute sulfuric acid at 145°C. Initial fermentations of the hydrolysate by strain FBR3 had lag times of approximately 30 h judged by ethanol production. Further experiments indicated that the acetate present in the hydrolysate could not solely account for the long lag. The lag phase was greatly reduced by growing the pre-seed and seed cultures on corn fiber hydrolysate. Ethanol yields for the optimized fermentations were 90% of theoretical. Maximum ethanol concentrations were 2.80% w/v, and the fermentations were completed in approximately 50 h. The optimal pH for the fermentation was 6.5. Below this pH, sugar consumption was incomplete and above it, excess base addition was required throughout the fermentation. Two alternative neutralization methods (overliming and overliming with sulfite addition) have been reported for improving the fermentability of lignocellulosic hydrolysates. These methods further reduced the lag phase of the fermentation, albeit by a minor amount.
Corn fiber, currently produced at wet milling facilities, is readily available as a potential fee... more Corn fiber, currently produced at wet milling facilities, is readily available as a potential feedstock for production of fermentable sugars. Destarched corn fiber (DSCF) can be conveniently prepared for enzymatic saccharification by treating with liquid hot-water. Treating DSCF with hot-water (HW-DSCF) at 160 • C for 20 min dissolved 58% of the solids and 75% of the xylan. Preparations of hydrolytic enzymes were next used to saccharify the cellulose and xylan. The needed enzymes were prepared from culture supernatants of Trichoderma reesei Rut C30 and Aspergillus niger NRRL 2001, each grown on HW-DSCF. The harvested cultures were found to have a broad range of carbohydrase activities. The enzyme profiles varied considerably from one another and the preparations were determined to be most effective for saccharifying HW-DSCF when used in combination. Monosaccharide sugar yields obtained using the blended preparations were 74 and 54% of the available arabinose and xylose, respectively. Arabinose and xylose yields were both further increased to 80% by increasing the hot-water pretreatment time to 30 min and adding a commercial preparation of -glucosidase, which also contained -xylosidase side-activity. Published by Elsevier Inc. (B.S. Dien). drates to fermentable monosaccharide sugars. Most research on corn fiber conversion to ethanol has focused on pretreating corn fiber with dilute sulfuric acid. Typically this involves lowering the pH to 1.0-1.3 with sulfuric acid and treating at a temperature of 150-180 • C for anywhere from 2 to 20 min . The sugar yields from these studies are generally very high . However, from a processing view-point, dilute-acid pretreatment has several shortcomings, including the need for expensive reactors capable of withstanding the combination of low-pH and high-temperature, formation of pretreatment-associated sideproducts that often stall the subsequent fermentation, and generation of acid-related waste streams (e.g., gypsum) that adds to the cost of waste treatment and complicates downstream processing.
... Fate of Bt Protein and Influence of Corn Hybrid on Ethanol Production BS Dien,1,2 RJ Bothast,... more ... Fate of Bt Protein and Influence of Corn Hybrid on Ethanol Production BS Dien,1,2 RJ Bothast,1 LB Iten,1 L. Barrios,3 and SR Eckhoff3 ... Ethanol produc-tion is expected to continue to increase as states phase out the use of methyl tertiary butyl ether (MTBE) as a fuel oxygenate. ...
Rice hulls, a complex lignocellulosic material with high lignin (15.38 ( 0.2%) and ash (18.71 ( 0... more Rice hulls, a complex lignocellulosic material with high lignin (15.38 ( 0.2%) and ash (18.71 ( 0.01%) content, contain 35.62 ( 0.12% cellulose and 11.96 ( 0.73% hemicellulose and has the potential to serve as a low-cost feedstock for production of ethanol. Dilute H 2 SO 4 pretreatments at varied temperature (120-190°C) and enzymatic saccharification (45°C, pH 5.0) were evaluated for conversion of rice hull cellulose and hemicellulose to monomeric sugars. The maximum yield of monomeric sugars from rice hulls (15%, w/v) by dilute H 2 SO 4 (1.0%, v/v) pretreatment and enzymatic saccharification (45°C, pH 5.0, 72 h) using cellulase, -glucosidase, xylanase, esterase, and Tween 20 was 287 ( 3 mg/g (60% yield based on total carbohydrate content). Under this condition, no furfural and hydroxymethyl furfural were produced. The yield of ethanol per L by the mixed sugar utilizing recombinant Escherichia coli strain FBR 5 from rice hull hydrolyzate containing 43.6 ( 3.0 g fermentable sugars (glucose, 18.2 ( 1.4 g; xylose, 21.4 ( 1.1 g; arabinose, 2.4 ( 0.3 g; galactose, 1.6 ( 0.2 g) was 18.7 ( 0.6 g (0.43 ( 0.02 g/g sugars obtained; 0.13 ( 0.01 g/g rice hulls) at pH 6.5 and 35°C. Detoxification of the acid-and enzyme-treated rice hull hydrolyzate by overliming (pH 10.5, 90°C, 30 min) reduced the time required for maximum ethanol production (17 ( 0.2 g from 42.0 ( 0.7 g sugars per L) by the E. coli strain from 64 to 39 h in the case of separate hydrolysis and fermentation and increased the maximum ethanol yield (per L) from 7.1 ( 2.3 g in 140 h to 9.1 ( 0.7 g in 112 h in the case of simultaneous saccharification and fermentation.
Alfalfa stems, reed canarygrass, and switchgrass; perennial herbaceous species that have potentia... more Alfalfa stems, reed canarygrass, and switchgrass; perennial herbaceous species that have potential as biomass energy crops in temperate regions; were evaluated for their bioconversion potential as energy crops. Each forage species was harvested at two or three maturity stages and analyzed for carbohydrates, lignin, protein, lipid, organic acids, and mineral composition. The biomass samples were also evaluated for sugar yields following pretreatment with dilute sulfuric followed by enzymatic saccharification using a commercial cellulase preparation. Total carbohydrate content of the plants varied from 518 to 655 g kg À1 dry matter (DM) and cellulose concentration from 209 to 322 g kg À1 DM. Carbohydrate and lignin contents were lower for samples from early maturity samples compared to samples from late maturity harvests. Several important trends were observed in regards to the efficiency of sugar recovery following treatments with dilute acid and cellulase. First, a significant amount of the available carbohydrates were in the form of soluble sugars and storage carbohydrates (4.3-16.3% wt/wt). Recovery of soluble sugars following dilute acid pretreatment was problematic, especially that of fructose. Fructose was found to be extremely labile to the dilute acid pretreatments. Second, the efficiency at which available glucose was recovered was inversely correlated to maturity and lignin content. However, total glucose yields were higher for the later maturities because of higher cellulose contents compared to the earlier maturity samples. Finally, cell wall polysaccharides, as determined by the widely applied detergent fiber system were found to be inaccurate. The detergent fiber method consistently overestimated cellulose and hemicellulose and underestimated lignin by substantial amounts. Published by Elsevier Ltd.
Clostridium beijerinckii P260 Agricultural residue hydrolysates Corn (Zea mays) stover Energy cro... more Clostridium beijerinckii P260 Agricultural residue hydrolysates Corn (Zea mays) stover Energy crop -switchgrass (Panicum virgatum) Fermentation Productivity Yield Overliming a b s t r a c t
In a series of experiments, untreated and ammonium hydroxide pretreated Klenow lowland variety sw... more In a series of experiments, untreated and ammonium hydroxide pretreated Klenow lowland variety switchgrasses are converted to reducing sugars using lowfrequency (20 kHz) ultrasound and commercially available cellulase enzyme. Results from experiments using untreated and pretreated switchgrasses with and without ultrasound are presented and discussed. In untreated switchgrass experiments, the combination of ultrasound and enzymes resulted in an increase of 7.5% in reducing sugars compared to experiments using just enzymes. In experiments using ammonium hydroxide pretreated switchgrass, the combination of ultrasound and enzymes resulted in an increase of 9.3% in reducing sugars compared to experiments using just enzymes. Experimental evidence indicates that there is a synergistic effect from the combination of ultrasound and enzymes which lowers the diffusion-limiting barrier to enzyme/substrate binding and results in an increase in reaction rate. Scanning electron microscopic images provide evidence that ultrasound-induced pitting increases substrate surface area and affects reaction rate and yield.
Switchgrass (Panicum virgatum L.) is a perennial C4 grass that is being developed as a bioenergy ... more Switchgrass (Panicum virgatum L.) is a perennial C4 grass that is being developed as a bioenergy crop because it has high production yields and suitable agronomic traits. Five switchgrass biomass samples from upland and lowland switchgrass ecotypes harvested at different stages or maturity were used in this study. Switchgrass samples contained 317.0-385.0 g glucans/kg switchgrass dry basis (db) and 579.3-660.2 g total structural carbohydrates/kg switchgrass, db. Carbohydrate contents were greater for the upland ecotype versus lowland ecotype and increased with harvest maturity. Pretreatment of switchgrass with dilute ammonium hydroxide (8% w/w ammonium loading) at 170 • C for 20 min was determined to be effective for preparing switchgrass for enzymatic conversion to monosaccharides; glucose recoveries were 66.9-90.5% and xylose recoveries 60.1-84.2% of maximum and decreased with increased maturity at harvest. Subsequently, pretreated switchgrass samples were converted to ethanol by simultaneous saccharification and fermentation using engineered xylose-fermenting Saccharomyces cerevisiae strain YRH400. Ethanol yields were 176.2-202.0 l/Mg of switchgrass (db) and followed a similar trend as observed for enzymatic sugar yields.
In a series of experiments, untreated and ammonium hydroxide pretreated Klenow lowland variety sw... more In a series of experiments, untreated and ammonium hydroxide pretreated Klenow lowland variety switchgrasses are converted to reducing sugars using low-frequency (20 kHz) ultrasound and commercially available cellulase enzyme. Results from experiments using untreated and pretreated switchgrasses with and without ultrasound are presented and discussed. In untreated switchgrass experiments, the combination of ultrasound and enzymes resulted in an increase of 7.5% in reducing sugars compared to experiments using just enzymes. In experiments using ammonium hydroxide pretreated switchgrass, the combination of ultrasound and enzymes resulted in an increase of 9.3% in reducing sugars compared to experiments using just enzymes. Experimental evidence indicates that there is a synergistic effect from the combination of ultrasound and enzymes which lowers the diffusion-limiting barrier to enzyme/substrate binding and results in an increase in reaction rate. Scanning electron microscopic images...
... Newman MM, Nguyen QA: Effects of temperature and moisture on dilute-acid steam explosion ... ... more ... Newman MM, Nguyen QA: Effects of temperature and moisture on dilute-acid steam explosion ... Dale BE, Ingram LO, Bothast RJ: Ethanol production from AFEX pretreated corn fiber ... Reshamwala S, Shawky BT, Dale BE: Ethanol production from enzymatic hydrolysates of AFEX ...
Use of agricultural residues for ethanol production requires pretreatment of the material to faci... more Use of agricultural residues for ethanol production requires pretreatment of the material to facilitate release of sugars. Physical–chemical pretreatment of lignocellulosic biomass can, however, give rise to side-products that may be toxic to fermenting microorganisms and hinder utilization of sugars obtained from biomass. Potentially problematic compounds include furan aldehydes formed by degradation of sugars, organic acids released from hemicellulose side-groups,
Journal of Industrial Microbiology and Biotechnology, 1999
We have developed a novel ethanologenic Escherichia coli strain FBR3 that is an efficient biocata... more We have developed a novel ethanologenic Escherichia coli strain FBR3 that is an efficient biocatalyst for converting mixed sugar streams (eg, arabinose, glucose, and xylose) into ethanol. In this report, the strain was tested for conversion of corn fiber hydrolysates into ethanol. Corn fiber hydrolysates with total sugar concentrations of 7.5% (w/v) were prepared by reacting corn fiber with dilute sulfuric acid at 145°C. Initial fermentations of the hydrolysate by strain FBR3 had lag times of approximately 30 h judged by ethanol production. Further experiments indicated that the acetate present in the hydrolysate could not solely account for the long lag. The lag phase was greatly reduced by growing the pre-seed and seed cultures on corn fiber hydrolysate. Ethanol yields for the optimized fermentations were 90% of theoretical. Maximum ethanol concentrations were 2.80% w/v, and the fermentations were completed in approximately 50 h. The optimal pH for the fermentation was 6.5. Below this pH, sugar consumption was incomplete and above it, excess base addition was required throughout the fermentation. Two alternative neutralization methods (overliming and overliming with sulfite addition) have been reported for improving the fermentability of lignocellulosic hydrolysates. These methods further reduced the lag phase of the fermentation, albeit by a minor amount.
Corn fiber, currently produced at wet milling facilities, is readily available as a potential fee... more Corn fiber, currently produced at wet milling facilities, is readily available as a potential feedstock for production of fermentable sugars. Destarched corn fiber (DSCF) can be conveniently prepared for enzymatic saccharification by treating with liquid hot-water. Treating DSCF with hot-water (HW-DSCF) at 160 • C for 20 min dissolved 58% of the solids and 75% of the xylan. Preparations of hydrolytic enzymes were next used to saccharify the cellulose and xylan. The needed enzymes were prepared from culture supernatants of Trichoderma reesei Rut C30 and Aspergillus niger NRRL 2001, each grown on HW-DSCF. The harvested cultures were found to have a broad range of carbohydrase activities. The enzyme profiles varied considerably from one another and the preparations were determined to be most effective for saccharifying HW-DSCF when used in combination. Monosaccharide sugar yields obtained using the blended preparations were 74 and 54% of the available arabinose and xylose, respectively. Arabinose and xylose yields were both further increased to 80% by increasing the hot-water pretreatment time to 30 min and adding a commercial preparation of -glucosidase, which also contained -xylosidase side-activity. Published by Elsevier Inc. (B.S. Dien). drates to fermentable monosaccharide sugars. Most research on corn fiber conversion to ethanol has focused on pretreating corn fiber with dilute sulfuric acid. Typically this involves lowering the pH to 1.0-1.3 with sulfuric acid and treating at a temperature of 150-180 • C for anywhere from 2 to 20 min . The sugar yields from these studies are generally very high . However, from a processing view-point, dilute-acid pretreatment has several shortcomings, including the need for expensive reactors capable of withstanding the combination of low-pH and high-temperature, formation of pretreatment-associated sideproducts that often stall the subsequent fermentation, and generation of acid-related waste streams (e.g., gypsum) that adds to the cost of waste treatment and complicates downstream processing.
... Fate of Bt Protein and Influence of Corn Hybrid on Ethanol Production BS Dien,1,2 RJ Bothast,... more ... Fate of Bt Protein and Influence of Corn Hybrid on Ethanol Production BS Dien,1,2 RJ Bothast,1 LB Iten,1 L. Barrios,3 and SR Eckhoff3 ... Ethanol produc-tion is expected to continue to increase as states phase out the use of methyl tertiary butyl ether (MTBE) as a fuel oxygenate. ...
Rice hulls, a complex lignocellulosic material with high lignin (15.38 ( 0.2%) and ash (18.71 ( 0... more Rice hulls, a complex lignocellulosic material with high lignin (15.38 ( 0.2%) and ash (18.71 ( 0.01%) content, contain 35.62 ( 0.12% cellulose and 11.96 ( 0.73% hemicellulose and has the potential to serve as a low-cost feedstock for production of ethanol. Dilute H 2 SO 4 pretreatments at varied temperature (120-190°C) and enzymatic saccharification (45°C, pH 5.0) were evaluated for conversion of rice hull cellulose and hemicellulose to monomeric sugars. The maximum yield of monomeric sugars from rice hulls (15%, w/v) by dilute H 2 SO 4 (1.0%, v/v) pretreatment and enzymatic saccharification (45°C, pH 5.0, 72 h) using cellulase, -glucosidase, xylanase, esterase, and Tween 20 was 287 ( 3 mg/g (60% yield based on total carbohydrate content). Under this condition, no furfural and hydroxymethyl furfural were produced. The yield of ethanol per L by the mixed sugar utilizing recombinant Escherichia coli strain FBR 5 from rice hull hydrolyzate containing 43.6 ( 3.0 g fermentable sugars (glucose, 18.2 ( 1.4 g; xylose, 21.4 ( 1.1 g; arabinose, 2.4 ( 0.3 g; galactose, 1.6 ( 0.2 g) was 18.7 ( 0.6 g (0.43 ( 0.02 g/g sugars obtained; 0.13 ( 0.01 g/g rice hulls) at pH 6.5 and 35°C. Detoxification of the acid-and enzyme-treated rice hull hydrolyzate by overliming (pH 10.5, 90°C, 30 min) reduced the time required for maximum ethanol production (17 ( 0.2 g from 42.0 ( 0.7 g sugars per L) by the E. coli strain from 64 to 39 h in the case of separate hydrolysis and fermentation and increased the maximum ethanol yield (per L) from 7.1 ( 2.3 g in 140 h to 9.1 ( 0.7 g in 112 h in the case of simultaneous saccharification and fermentation.
Alfalfa stems, reed canarygrass, and switchgrass; perennial herbaceous species that have potentia... more Alfalfa stems, reed canarygrass, and switchgrass; perennial herbaceous species that have potential as biomass energy crops in temperate regions; were evaluated for their bioconversion potential as energy crops. Each forage species was harvested at two or three maturity stages and analyzed for carbohydrates, lignin, protein, lipid, organic acids, and mineral composition. The biomass samples were also evaluated for sugar yields following pretreatment with dilute sulfuric followed by enzymatic saccharification using a commercial cellulase preparation. Total carbohydrate content of the plants varied from 518 to 655 g kg À1 dry matter (DM) and cellulose concentration from 209 to 322 g kg À1 DM. Carbohydrate and lignin contents were lower for samples from early maturity samples compared to samples from late maturity harvests. Several important trends were observed in regards to the efficiency of sugar recovery following treatments with dilute acid and cellulase. First, a significant amount of the available carbohydrates were in the form of soluble sugars and storage carbohydrates (4.3-16.3% wt/wt). Recovery of soluble sugars following dilute acid pretreatment was problematic, especially that of fructose. Fructose was found to be extremely labile to the dilute acid pretreatments. Second, the efficiency at which available glucose was recovered was inversely correlated to maturity and lignin content. However, total glucose yields were higher for the later maturities because of higher cellulose contents compared to the earlier maturity samples. Finally, cell wall polysaccharides, as determined by the widely applied detergent fiber system were found to be inaccurate. The detergent fiber method consistently overestimated cellulose and hemicellulose and underestimated lignin by substantial amounts. Published by Elsevier Ltd.
Clostridium beijerinckii P260 Agricultural residue hydrolysates Corn (Zea mays) stover Energy cro... more Clostridium beijerinckii P260 Agricultural residue hydrolysates Corn (Zea mays) stover Energy crop -switchgrass (Panicum virgatum) Fermentation Productivity Yield Overliming a b s t r a c t
In a series of experiments, untreated and ammonium hydroxide pretreated Klenow lowland variety sw... more In a series of experiments, untreated and ammonium hydroxide pretreated Klenow lowland variety switchgrasses are converted to reducing sugars using lowfrequency (20 kHz) ultrasound and commercially available cellulase enzyme. Results from experiments using untreated and pretreated switchgrasses with and without ultrasound are presented and discussed. In untreated switchgrass experiments, the combination of ultrasound and enzymes resulted in an increase of 7.5% in reducing sugars compared to experiments using just enzymes. In experiments using ammonium hydroxide pretreated switchgrass, the combination of ultrasound and enzymes resulted in an increase of 9.3% in reducing sugars compared to experiments using just enzymes. Experimental evidence indicates that there is a synergistic effect from the combination of ultrasound and enzymes which lowers the diffusion-limiting barrier to enzyme/substrate binding and results in an increase in reaction rate. Scanning electron microscopic images provide evidence that ultrasound-induced pitting increases substrate surface area and affects reaction rate and yield.
Switchgrass (Panicum virgatum L.) is a perennial C4 grass that is being developed as a bioenergy ... more Switchgrass (Panicum virgatum L.) is a perennial C4 grass that is being developed as a bioenergy crop because it has high production yields and suitable agronomic traits. Five switchgrass biomass samples from upland and lowland switchgrass ecotypes harvested at different stages or maturity were used in this study. Switchgrass samples contained 317.0-385.0 g glucans/kg switchgrass dry basis (db) and 579.3-660.2 g total structural carbohydrates/kg switchgrass, db. Carbohydrate contents were greater for the upland ecotype versus lowland ecotype and increased with harvest maturity. Pretreatment of switchgrass with dilute ammonium hydroxide (8% w/w ammonium loading) at 170 • C for 20 min was determined to be effective for preparing switchgrass for enzymatic conversion to monosaccharides; glucose recoveries were 66.9-90.5% and xylose recoveries 60.1-84.2% of maximum and decreased with increased maturity at harvest. Subsequently, pretreated switchgrass samples were converted to ethanol by simultaneous saccharification and fermentation using engineered xylose-fermenting Saccharomyces cerevisiae strain YRH400. Ethanol yields were 176.2-202.0 l/Mg of switchgrass (db) and followed a similar trend as observed for enzymatic sugar yields.
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